Tuberculosis (TB) is the leading infectious killer of youth and adults and the first most common infectious disease worldwide. One third of the world's population is currently infected and 20 million of those infected are active cases; TB will kill 30 million people this decade. More than 50 million people may already be infected with multidrug-resistant (MDR) strains of TB. Drug resistance has been brought about because of complacency in the public health sector and poorly managed TB control programs. Prior to MDR tuberculosis, the success rate of drug combination treatment was greater than 90%, even in AIDS patients. MDR tuberculosis, however, is not only highly infectious but also essentially incurable with a mortality of 50%. TB is now becoming the leading cause of death among HIV positive people where it kills much more rapidly with a fatality of 80%.
Tuberculosis is caused by infection with Mycobacterium tuberculosis, a bacillus bacterium. It is spread by aerosol droplets and causes irreversible lung destruction. If it escapes the lung it may cause systemic disease affecting many organs including bones, joints, liver, spleen, gastrointestinal tract and brain. 50% of people exposed to M. tuberculosis are infected with the bacteria and 15% of those infected develop disease. Poverty, malnutrition and overpopulation contribute dramatically to the perseverance and wild spread of tuberculosis.
Past means of controlling TB have involved the use of combinations of antibiotics. Recently, because of complications due to multidrug-resistant strains, the number and combination of antibiotics administered must be individually tailored depending on the strain the patient is harboring. In extreme cases, surgical removal of the infected portion of the lung is required.
Traditionally, the diagnosis of TB has been made on the basis of clinical findings and chest radiographs and confirmed by sputum or tissue smears that show TB bacilli. These methods remain the “gold standard” for diagnosis, but development of DNA probes, polymerase chain reaction (PCR) assays, and liquid media now allow more sensitive and rapid diagnosis. Unfortunately, increased sensitivity of rapid techniques is not always associated with increased specificity.
Skin testing should be used in conjunction with other clinical findings and is not a sensitive or specific test for establishing the diagnosis when the patient had been vaccinated with BCG or infected by Mycobacterium other than tuberculosis. In extrapulmonary TB, site-specific tissue or fluid samples or both are submitted for smear, culture, and histological analysis. Typically, histological features of a tuberculous lesion include caseating and noncaseating granulomata with giant cells. Rapid laboratory testing to identify and determine the drug susceptibility of M. tuberculosis isolates is vital to effective diagnosis, treatment, and control of TB in the community.
Clearly, expedient diagnosis is important in controlling the spread of tuberculosis. Sputum samples, evaluated first by direct microscopic evaluation (smear), are visualized with either the easily detected acid-fast fluorochrome dye auramine O, or the more specific Ziehl-Neelsen stain. Specimens are cultured on either solid media (Lowenstein-Jensen slant), or are grown in a liquid medium, such as the BACTEC automated radiometric system. Next, biochemical or nucleic acid probe testing is used to identify various strains. Isolates are tested for resistance to commonly used antituberculosis drugs, often by using the new method of susceptibility testing in liquid broth rather than the traditional agar dilution method.
The Ziehl-Neelsen carbolfuchsin or Kinyoun carbolfuchsin stains have been essential in TB diagnosis for nearly 100 years. Although less sensitive than culture, the acid-fast smear is a rapid and inexpensive test that can be performed with a minimum of equipment and is very specific for mycobacteria. Depending on the bacterial load, a single sputum smear has sensitivity between 22% and 80%, but the yield is improved when multiple sputum specimens are examined.
Most laboratories in the United States use fluorochrome stains, such as auramine-rhodamine stain. With these techniques, mycobacteria fluoresce with a bright orange color and can be easily seen on low-power microscopy, increasing the sensitivity of the smears.
The Amplified Mycobacterium Tuberculosis Direct Test (Gen-Probe) targets mycobacterial ribosomal RNA by transcription-mediated amplification. The test uses DNA probes that are highly specific for M. tuberculosis species. It is best used (and only approved for use) in patients in whom acid-fast bacilli smears are positive and cultures are in process. Since specificity is less than 100%, even in patients with positive smears, occasional false-positive results do occur, usually in patients with nontuberculous mycobacterial infections.
This technique amplifies even very small portions of a predetermined target region of M. tuberculosis-complex DNA. The test uses an automated system that can rapidly detect as few as one organism from sputum, bronchoalveolar lavage, blood, cerebrospinal fluid, pleural fluid, or other fluid and tissue samples and has shown sensitivity and specificity of nearly 90% in pulmonary disease.
The Mantoux test is the preferred and standard skin test for detecting TB. It involves injection of 5 TU of purified protein derivative (PPD, tuberculin), usually 0.1 mL, intradermally. Induration is then assessed at 48 to 72 hours. The extent of induration (not erythema) should be measured across two diameters at right angles and the two measurements then averaged. Interobserver variability can be decreased through the use of a ballpoint pen carefully brought from outside the zone of induration toward the center. As the area of induration is reached, resistance increases, and marking should stop at the outer edge of induration. However, about 20% of patients with active TB may have negative skin tests, and some populations have an even higher incidence of false-negative results. For example, false-negative rates up to 50% have been reported in patients with advanced HIV infection. Alternately, false-positive results may occur in patients infected by other nontuberculous mycobacteria (e.g., Mycobacterium avium complex). Therefore, a negative skin test never rules out TB, and a positive skin test alone does not establish the diagnosis.
The US Centers for Disease Control and Prevention and the World Health Organization recommend initial susceptibility testing for all M. tuberculosis isolates because of the emergence of drug resistance worldwide.
This ingenious assay uses the fluorescent capabilities of fireflies genetically implanted in M. tuberculosis. The procedure offers the possibility of testing mycobacterial drug susceptibility in hours. It is in the development stages but may become widely available in the next few years.
Given the above, current available assays cannot quickly and completely detect M. tuberculosis. It requires a quick assay with high specificity and sensitivity to detect M. tuberculosis from available samples.